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Fig. 5 | Molecular Neurodegeneration

Fig. 5

From: Upregulation of Ca2+-binding proteins contributes to VTA dopamine neuron survival in the early phases of Alzheimer’s disease in Tg2576 mice

Fig. 5

Overexpression of CB and CR in DA neurons of the Tg2576 VTA. A. Representative confocal image of coronal midbrain section showing TH+ (in red) together with Calbindin (CB+) or Calretinin (CR+) (both in green; scale: 100 μm). B. Stereological quantification of TH+ neurons co-expressing Ca2+-binding proteins (CB or CR) in the VTA of WT and Tg2576 mice of 3- and 6- months of age. TH+CB+ (3 months: WT: n = 6; Tg2576: n = 6; 6 months: WT: n = 4; Tg2576: n = 5 mice. Two-way ANOVA for Genotype vs Age: Interaction: F1,17 = 2.343, p = 0.1443; Age: F1,17 = 1.625, p = 0.2196; Genotype: F1,17 = 20.72, p = 0.0003. WT 3 m vs Tg2576 3 m: Unpaired t-test: *p = 0.0447; WT 6 m vs Tg2576 6 m: Unpaired t-test: **p = 0.0048; Tg2576 3 m vs Tg2576 6 m: Unpaired t-test: **p = 0.0067). TH+CR+ (3 months: WT: n = 4; Tg2576: n = 5; 6 months: WT: n = 5; Tg2576: n = 7 mice. Two-way ANOVA for Genotype vs Age: Interaction: F1,17 = 0.834, p = 0.3714; Genotype: F1,17 = 22.66, p = 0.0002; Age: F1,17 = 22.6, p = 0.0002; WT 3 m vs Tg2576 3 m: *p = 0.0352; WT 6 m vs Tg2576 6 m: **p = 0.0017; Tg2576 3 m vs Tg2576 6 m: ***p = 0.0005). C. Representative western blots from the midbrain of 1-, 3- and 6-month-old WT and Tg2576 mice, and plots showing the levels (expressed as % of WT) of CB (left panel) or CR (right panel) normalized to Actin as loading control. CB: 1 month: n = 5 mice per genotype; 3 months: WT: n = 9; Tg2576: n = 7 mice; Unpaired t-test: *p = 0.0156; 6-months: WT: n = 12; Tg2576: n = 10 mice. CR: 1 month: n = 5 mice per genotype; 3 months: WT: n = 8; Tg2576: n = 9; 6-months: WT: n = 10; Tg2576: n = 11; Unpaired t-test: *p = 0.0228. D-E. Panels show the analysis of confocal Z-stack double immunofluorescent labelling for TH (blue) and CB (D) or CR (E; both in green) in the VTA of coronal sections from 1-, 3- and 6-months-old WT and Tg2576 mice (scale: 20 μm). Box and whisker plots show the densitometric analysis of CB and CR levels in VTA TH+ neurons (D. 1 months-old: WT n = 6 mice, Tg2576 n = 5 mice; 3 months-old: n = 6 mice per genotype; Welch’s: **p = 0.0046; 6 months-old: WT n = 6 mice, Tg2576 n = 5 mice. E. 1 months-old: n = 4 mice per genotype; 3 months-old: WT n = 5 mice, Tg2576 n = 4 mice; Unpaired t-test: *p = 0. 0263. 6 months-old: WT n = 6 mice, Tg2576 n = 5 mice). F. The plot shows the amount of free [Ca2+] expressed as 340/380 ratio of Fura-2 in the PN o 3-month-old WT and Tg2576 mice (WT: n = 9 neurons,3 mice; Tg2576: n = 10 neurons, 3 mice; Welch’s t-test, **p = 0.0024). G. Representative confocal images of TUNEL-negative (TUNEL-) and -positive (TUNEL+) dopaminergic neurons expressing CB from the VTA of 3-month-old Tg2576 mice (scale bar: 10 μm). The graph shows densitometric levels of CB in TUNEL- and TUNEL+ neurons (TUNEL-: n = 19 neurons; 3 mice; TUNEL+: n = 16 neurons; 3 mice; Unpaired t-test: *p = 0.0141)

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